Proliferation and regeneration of retrogradely labeled adult rat corticospinal neurons in culture.

These are the first studies to demonstrate that adult rat corticospinal tract (CST) neurons, which were identified by retrograde neuronal labeling, retain regenerative and proliferative potential. To determine if adult CST neurons undergo cell division, we tested if these retrogradely labeled cells synthesize DNA by adding BrdU to the cultures 24 h prior to fixation of the cells. The result shows that adult corticospinal tract neurons are capable of DNA synthesis, and our total cell counts with labeled cells counts further suggest that these cells undergo cell division.

Transcranial Doppler ultrasonography-guided management of internal carotid artery closure.

OBJECTIVE: To emphasize the integrated use of transcranial Doppler ultrasonography (TCD) in the management of internal carotid artery (ICA) closure. METHODS: Thirty-three patients being considered for ICA closure underwent TCD assessment, vasomotor reserve testing/estimation, and carotid artery test occlusion with concomitant middle cerebral artery (MCA) blood velocity (V(MCA)) monitoring, including calculation of the MCA pulsatility index. Twelve of these patients proceeded to undergo ICA sacrifice. Sequential TCD sonograms guided their postoperative treatment. RESULTS: ICA aneurysms and neck neoplasms affected the TCD results and vasomotor reserve insignificantly, whereas carotid-cavernous fistulae induced characteristic circulatory alterations. The 10 subjects who tolerated ICA sacrifice hemodynamically all showed an initial decrease in the ipsilateral V(MCA) to > or =60% of the preocclusion value and a progressively decreasing MCA pulsatility index during carotid artery test occlusion. The two patients who developed hemodynamic cerebral infarctions exhibited a decrease in V(MCA) to <60% and a MCA pulsatility index that remained stable after a vast initial reduction. Postoperative hypervolemic and hypertensive support was safely titrated in all patients who received postoperative TCD surveillance, providing an ipsilateral V(MCA) of > or =80% of the preocclusion value. ICA closure permanently altered the cerebral circulatory pattern. CONCLUSION: The hemodynamic outcome of ICA sacrifice can be correctly predicted by using the TCD occlusion test. TCD provides the means to titrate the extent of postoperative hypervolemic/hypertensive support.

Surgical treatment of anterior circulation aneurysms.

The purpose of this paper is to present the results, assessed by an independent observer, of surgical treatment of 428 consecutive patients harbouring aneurysms of the anterior circulation, together with a review of relevant anatomy and operative strategy. At follow-up (mean 5.6 years) 89.3% lived at home and were independent, 5.1% lived at home but needed some kind of assistance, 2.0% lived in institution, whereas information was unavailable in 3.6% of living patients. Two hundred and fifty-three patients (64.5%) had unchanged employment status, 0.3% worked in sheltered environment, whereas 30.9% went out of work due to their subarachnoid hemorrhage (SAH). Information about employment status was unavailable in 4.3%. For aneurysms of the internal carotid, anterior communicating and middle cerebral artery, respectively, mortality was 3.2, 3.9 and 5.6%, whereas 92.0, 88.1 and 89.0% of surviving patients lived at home and were independent and 67.0, 63.6 and 63.0% had unchanged employment status. Three-months mortality of all causes was 4.2%. In the postoperative period 53 (12.4%) patients developed clinical signs of vasospasms, 6 (1.4%) had cardiac infarction, 4 (0.9%) lung oedema, 4 (0.9%) deep vein thrombosis, and 7 patients (1.6%) infection. During the follow-up period shunt-dependent hydrocephalus developed in 4.2% and 0.2% had a subsequent SAH from the same aneurysm. Forty-three patients were on anticonvulsive therapy.

Varying impact of common carotid artery digital compression and internal carotid artery balloon test occlusion on cerebral hemodynamics.

BACKGROUND: The purpose of this study was to compare the cerebral hemodynamic changes brought about by common carotid artery (CCA) digital compression and angiographic internal carotid artery (ICA) balloon occlusion. METHODS: Bilateral transcranial Doppler ultrasonographic monitoring of the middle cerebral artery blood velocity (VMCA) was performed in 12 subjects with neck neoplasms or traumatic carotid-cavernous fistulas. The MCA pulsatility index (PIMCA) and hemodynamic tension (Uhem MCA) were calculated. RESULTS: Common carotid artery compression provoked the largest drop in ipsilateral VMCA, PIMCA, and Uhem MCA. Common carotid artery compression caused a steal of blood from the intra- to the extracranial circulation, with the discrepancy in hemodynamic findings between CCA and ICA test occlusions being dependent on the quantity of reversed ipsilateral ICA blood flow. CONCLUSION: If the carotid artery is to be sacrificed, permanent ICA closure is the procedure of choice with respect to the occurrence of cerebral ischemic lesions in patients with neck neoplasms and ICA flow reversal during CCA compression.

Cerebral haemodynamics in internal carotid artery trial occlusion.

The purpose of this study was to analyse the cerebral haemodynamic changes brought about by trial occlusion of the internal carotid artery (ICA). Sixteen patients with surgically inaccessible cerebral aneurysms, carotid cavernous fistulas or neck neoplasms were monitored with transcranial Doppler ultrasonography (TCD) during 90-120 s angiographic ICA balloon occlusion or ICA closure with a Selverstone clamp. The blood velocity (V) was registered continuously in both middle cerebral arteries (MCA) while the pulsatility index (PIMCA) and haemodynamic tension (Uhem MCA) were calculated. ICA closure led to an instantaneous drop in the ipsilateral VMCA, PIMCA and Uhem MCA. The VMCA thereafter increased gradually until reaching a stable level. The subjects were grouped into those with initial drops in VMCA to > or = 60% of pre-occlusion value (group 1) and those that fell to < 60% (group 2), respectively. In group 1 autoregulatory mechanisms made the PIMCA decline further, while the Uhem MCA remained unaltered during ICA closure. In group 2, however, the PIMCA did not change further, while the Uhem MCA increased slightly. The cerebral haemodynamic features during ICA test occlusion were thus essentially different in the two groups. On re-opening the ICA, there was an overshoot in VMCA and Uhem MCA. Contralaterally, the VMCA was increased during ICA occlusion. Seven of the patients later had their ICA closed permanently. While none of five group 1 patients developed haemodynamic complications, two group 2 individuals experienced haemodynamic stroke. Assuming ICA sacrifice is feasible when test occlusion results in an ipsilateral initial reduction in VMCA to > or = 60% of pre-occlusion value, the corresponding limit for the Uhem MCA is > or = 40%. In the pre-operative evaluation of the haemodynamic risk related to ICA loss, TCD emerges as a reliable method. It also seems to allow for the reduction of test occlusion time to 90-120 s.

Overexpression of E2F1 in astrocytes leads to neoplastic transformation and changes in expression of retinoblastoma family members.

Overexpression of mouse E2F1 full-length but not truncated forms results in neoplastic transformation of astrocytes in vitro. This neoplastic transformation is accompanied with changes in cell morphology and expression of cell cycle regulators. Transformed astrocytes have higher expression of cdk2, pRb, and p107 than control astrocytes. However, expression of glial fibrillary acidic protein (GFAP) and p130 is reduced in transformed astrocytes.

Cell type specific regulation of COUP-TF II promoter activity.

COUP-TF family orphan receptors regulate activity of ligand-activated nuclear hormone receptors or function independently in the regulation of gene expression. COUP-TF II has a complex expression pattern suggesting that different mechanisms are involved in the regulation of its expression. We isolated the 5' regulatory region of the mouse COUP-TF II gene and demonstrated that the basal promoter is localized in a -200 bp region 5' from the transcription start site. All-trans retinoic acid and dibutyryl cyclic AMP have cell type specific effects on COUP-TF II promoter activity. The effect of cyclic AMP is mediated by the cyclic AMP response element that is localized 74 nucleotides upstream from the major transcriptional start. In vitro promoter analyses also demonstrated that the effect of all-trans RA is not directly mediated by the binding of RARs or RXRs to the promoter sequence.

All-trans retinoic acid affects the expression of orphan receptors COUP-TF I and COUP-TF II in the developing neural tube.

The effect of all-trans retinoic acid (RA) on chicken ovalbumin upstream promoter-transcription factor (COUP-TF) I and COUP-TF II expression in the developing cervical spinal cord and telencephalon was examined using embryonic day 11 and 13 mice. All-trans RA treatment results in changes in expression of COUP-TF I and COUP-TF II genes in the spinal cord and telencephalon. COUP-TF I mRNA levels were reduced in the spinal cord ventricular zone (VZ) after treatment, whereas in the telencephalon, mRNA levels were unaffected in the VZ and increased in the intermediate zone (IZ). COUP-TF II mRNA levels were increased in the spinal cord VZ after treatment, while in the telencephalon, IZ mRNA levels were decreased in the E11 embryos and increased in E13 embryos.

Cerebral haemodynamic considerations in obstructive carotid artery disease.

46 subjects with obstructive carotid artery disease were investigated with transcranial Doppler ultrasonography. Their baseline blood velocities (V) in the middle, anterior and posterior cerebral artery (MCA, ACA and PCA) and in the extracranial internal carotid artery (ICA) were measured and the pulsatility index (PI) calculated for each vessel. Thereafter the vasomotor reserve in both MCAs was tested. Typical patterns of V, PI and vasomotor reactivity are presented. Arterial collaterals were recognized by their relatively increased velocities. We demonstrated a close association of the baseline variables V and PI and the total vasomotor reactivity (hypocapnic plus no, hypercapnic response) by calculating an index of Uhem related to the cerebrovascular tone. The Uhem index is expressed by: Uhem index = VMCA.PIMCA/VPCA.PIPCA The relationship between Uhem index and the total vasomotor reactivity seemed to correspond to a hyperbolic curve. The hyperbolic tangent of Uhem index and total vasomotor reactivity correlated highly significantly, r = 0.8203, p < 0.0001, n = 49, the best fit for the regression line was Y = -0.005 + Uhem index 51.3. On the 99% significance level an Uhem index > or = 0.94 indicated normal total cerebral vasomotor reactivity in contrast to an impaired reactivity when < or = 0.81. Findings in 20 patients investigated post hoc supported the validity of our concept.

Haemodynamic classification of symptomatic obstructive carotid artery disease.

63 subjects with symptomatic obstructive carotid artery disease were investigated with transcranial Doppler ultrasonography. Their blood velocities at rest (V) in the middle and posterior cerebral artery (MCA and PCA) and in the extracranial internal carotid artery were measured and the pulsatility index (PI) and Uhem index (VMCA.PIMCA/VPCA.PIPCA) calculated. The vasomotor responses in both MCAs were also tested. The subjects were divided into groups based on the findings on physical examination and cerebral computed tomography. In the patient group with lacunar/territorial infarction we found in the stroke hemisphere: VMCA > VPCA, PIMCA = PIPCA and normal values for the Uhem index and total vasomotor reactivity. In the patient group with watershed infarction this hemisphere was characterized by: VMCA < VPCA, PIMCA < PIPCA and subnormal scores for the Uhem index and total vasomotor reactivity. Displaying features from both stroke groups, we obtained in the hemisphere of interest in patients with transient ischaemic attacks: VMCA = VPCA, PIMCA < PIPCA and normal values for the Uhem index and total vasomotor reactivity. Five patients with clinical evidence of stroke but with negative cerebral computed tomography findings had scores similar to those of the watershed group of patients. For the stroke patients, individual measurements of V, PI and total vasomotor reactivity failed to clearly identify to which stroke group a subject might belong. However, such an identification was achieved in all subjects when using the Uhem index. The Uhem index data in patients with transient ischaemic attacks suggest two subgroups with different pathogenesis underlying, the ischaemic events.

Helix-loop-helix transcription factors regulate Id2 gene promoter activity.

Id-like helix-loop-helix (HLH) transcription factors are involved in the regulation of proliferation and differentiation of several cell types. We isolated 5' regulatory region of mouse Id2 gene and demonstrated that it contains several E-box clusters. These E-boxes mediate stimulatory effects of basic-HLH (bHLH) transcription factors ME1, ME2, and NSCL1 on Id2 promoter activity. Co-expression of Id2 blocks the stimulatory effect of bHLH transcription factors which suggests the presence of feedback loops in Id2 transcriptional regulation. Overexpression of NSCL1 in F9 cells blocks the downregulation of Id2 gene expression during retinoic acid induced differentiation. Our data demonstrate that bHLH transcription factors regulate Id2 gene expression.

Expression screening reveals an orphan receptor chick ovalbumin upstream promoter transcription factor I as a regulator of neurite/substrate-cell contacts and cell aggregation.

A rat homologue of chick ovalbumin upstream promoter transcription factor I (COUP-TF I) was isolated using an expression cloning method developed to isolate neurite outgrowth inhibitors. Overexpression of COUP-TF I in 3T3 fibroblasts resulted in reduction of stable contact formation between neurites and transfected cells. Additionally, COUP-TF I enhanced retinoic acid response element-dependent reporter gene expression in 3T3 fibroblasts, indicating that COUP-TF I can modulate transcriptional activation in these cells. Our data suggest that COUP-TF transcription factors are involved in the regulation of cell surface molecules during neruogenesis.

Cyclin E is expressed in neurons and forms complexes with cdk5.

Expression of cyclin dependent kinase cdk5 and its regulator p35 has been shown in the cytoplasm of adult neurons. Here we demonstrate that another potential regulator of cdk5, cyclin E, is expressed in the nervous system and forms complexes with cdk5. Western blot analyses identifies expression of two forms of cyclin E in the mouse nervous system with the 56 kDa form mainly expressed in neurons and 51 kDa form expressed in astrocytes and oligodendrocytes.

Orphan receptor COUP-TF I antagonizes retinoic acid-induced neuronal differentiation.

Chicken ovalbumin upstream promoter-transcription factors (COUP-TF) are expressed in the developing nervous system and interact with nuclear hormone receptors to regulate expression of different genes. The role of COUP-TF orphan receptors in neurogenesis is virtually unknown. To study the possible function of COUP-TF I during neuronal differentiation, we generated COUP-TF I overexpressing teratocarcinoma PCC7 cell lines and analyzed retinoic acid (RA)-induced neuronal differentiation of these cells. COUP-TF I overexpression results in the blockade of morphological differentiation after induction to differentiate. COUP-TF I represses expression of microtubule-associated protein 2 (MAP2) gene and delays induction of growth-associated protein 43 (GAP43) gene expression. In contrast, expression of the neurofilament light subunit (NF-L) gene is not affected by COUP-TF I overexpression during neuronal differentiation. Also, cells overexpressing COUP-TF I do not stop proliferating after RA and dBcAMP treatment and possess suppressed transcriptional activation from different RA response elements. These results suggest that COUP-TF I plays an important role in regulating RA-induced neuronal differentiation.

Expression of Rb, E2F1, cdc2, and D, and B cyclins in developing spinal cord.

The spatial and temporal distribution of transcripts for the tumor suppressor gene Rb, transcription factor E2F1, cdc2 kinase, cyclins D1, D2, B1 and B2 during neurogenesis of the spinal cord was determined by in situ hybridization. The Rb and E2F1 transcripts were detectable in proliferating and differentiating cells. By contrast, cdc2, cyclins D1, B1 and B2 are expressed in the ventricular zone where proliferating cells are localized. Cyclin D2 mRNA was detectable only in the marginal zone of the developing neural tube. Electrophoretic mobility shift analyses demonstrated a changing pattern of DNA/protein complexes that bind to E2F binding site. These observations suggest that Rb and E2F1 may be involved in the early stages of neuronal differentiation in addition to the cell cycle regulation.

Isolation of genes which block neuronal differentiation of teratocarcinoma PCC7 cells.

Since several viral oncoproteins block differentiation and induce proliferation of differentiated cells, we developed an expression screening method to isolate cDNAs which block neuronal differentiation and induce proliferation of teratocarcinoma cells. Mouse E2F1, RNP-1, and RNP-2 (Regulator of Neuronal Proliferation) were isolated using the developed screening method. Overexpression of E2F1, RNP-1, and RNP-2 cDNAs in neuronally differentiated teratocarcinoma PCC7 cells results in blocking differentiation and initiation of proliferation. Also, expression of RNP-1 and RNP-2 blocks the expression of neurofilament-L and GAP-43 genes in PCC7 cells.

DNA synthesis is induced in adult neurons after expression of E2F1 and E1A.

Differentiated neurons in several brain regions express the retinoblastoma (Rb) tumor suppressor gene. Changing the tumor suppressor function of Rb by expressing transcription factor E2F1 and viral oncoprotein E1A in cerebellar granular neurons in vitro and in cerebral cortical neurons in vivo results in the induction of DNA synthesis in these neurons. Immunoliposome-mediated transfection of E1A and E2F1 cDNAs into the adult cortical neurons of rats in vivo results in initiation of DNA synthesis in 5-15% of the transfected neurons. These results indicate that expression of Rb may be necessary to prevent induction of differentiated neurons to proliferate since many mitogenic growth factors are expressed in the brain.

Class A basic helix-loop-helix transcription factors in early stages of chick neural tube development: evidence for functional redundancy.

Basic Helix-Loop-Helix (bHLH) transcription factors play essential role in differentiation of several cell types including neurons. We isolated chick bHLH transcription factor E12 cDNA and demonstrated its expression in developing neural tube interneurons. To examine the function of class A bHLH transcription factors we blocked their expression using antisense oligonucleotides. Simultaneous blocking of class A bHLH transcription factors in cultured neural tube cells results in the reduction of differentiating neurons. Blocking the expression of individual class A bHLH transcription factors has no detectable effect. These results demonstrate that class A bHLH transcription factors have a functional redundancy during neuronal development.

Superselective angiography, embolisation and surgery in treatment of arteriovenous malformations of the brain.

We prospectively evaluated a diagnostic and therapeutic procedure combining superselective angiography, endovascular embolisation and surgery in arteriovenous malformations (AVM) of the brain. Our aim was to achieve the best possible result exposing the patients to the lowest possible risk. During a 3 year period 48 patients were studied, of whom 22 (46%) were treated by incomplete preoperative embolisation and subsequent complete surgical removal of the AVM. All angiography and embolisations were performed with the Tracker catheter system using polyvinyl alcohol particles and/or platinum fibre coils. The other 26 patients were treated as follows: incomplete embolisation in 11 (23%), complete embolisation in 2 (4%), angiography and surgery in 3 (6%) and combined embolisation and radiation in 7 (15%); 3 patients were excluded from any therapy after superselective angiography. The total number of acute complications and permanent sequelae were 8 (17%) and 3 (6%) respectively. We concluded that superselective cerebral angiography, a relatively safe procedure, should play an important role in deciding treatment strategies for AVMs of the brain. The risks of combined embolisation and surgery are probably lower than those of either alone. In a few cases surgery or embolisation alone carries an unequivocally lower risk, and combined therapy is not justifiable.

Effects of cultured adrenal chromaffin cell implants on hindlimb reflexes of the 6-OHDA lesioned rat.

The effects of implantation of cultured adrenal medullary cells on the recovery of neurotransmitter specific reflex activity were studied in the rat spinal cord using electrophysiological testing methods. Cell suspensions of cultured neonatal adrenal medullary chromaffin (AM) cells (which produce catecholamines), or Schwann (Sc) cells (controls) were implanted into the lumbar region of the spinal cord 2 weeks after catecholamine (CA) denervation by intracisternal injection of 6-hydroxydopamine (6-OHDA). All cells were taken from 7 day neonates and cultured for 10 days in the presence of nerve growth factor (NGF). Three months after implantation, the extent of implant-associated recovery of reflex activity was determined by measuring electromyogram (EMG) activity and force associated with the long latency component of the hindlimb withdrawal reflex (which is CA modulated). After the electrophysiological testing, rats were anesthetized, and the spinal cords were rapidly removed and frozen. Spinal cords were sectioned longitudinally, and implanted cells were visualized using glyoxylic acid techniques. Labelled sections were examined to determine cell survival. Results indicate that 1) chromaffin cells survive for 3 months in the segments of the cord into which they have been implanted and 2) rats implanted with AM cells have significantly more forceful withdrawal reflexes than those that received Sc cells or received no implant after lesioning.